The Rev protein is absolutely essential for the production of the viral structural proteins and thus inhibition of Rev function leads to inhibition of HIV replication through prevention of structural protein synthesis and the assembly of HIV particles (For a review on Rev see Pollard, V. W., and M. H. Malim. 1998. Annu Rev Microbiol. 52:491-532). This is most easily shown using proviral clones that lack a functional rev gene, in that they have no replicative ability (Feinberg et al 1986 Cell 46:807-817; Bray et al. 1994 PNAS 91:1256-60). In the absence of functional Rev, genomic HIV RNA and several other HIV mRNAs remain the cell nucleus (Felber et al. 1989 Proc Natl Acad Sci USA. 86:1496-1499; Hammarskjold et al. 1989, J Virol. 63:1959-1966; Malim et al. Nature. 338:254-257). Since the RNA is not exported to the cytoplasm, viral structural proteins are not made and the infectious cycle cannot continue. Thus, it is clear that modalities inhibiting the function of Rev can form the basis for therapy against HIV infection and AIDS.
Although the Rev/RRE export pathway is still not fully understood, several important steps in this pathway have now been identified. The current understanding of this process is schematically shown in FIG. 1. The pathway starts with the import of Rev into the nucleus (step 1). In step 2, Rev binds specifically to RNA containing a specific RNA element called the Rev Response Element (RRE). In step 3, Rev multimerizes on the RRE in a process believed to involve protein-protein as well as protein-RNA interactions. In step 4, the Rev-RRE complex is recognized by CRM1 and RAN-GTP, which initiates the export process and eventually targets the complex to the nuclear pore (Step 5), where it interacts with nucleoporins resulting in translocation to the cytoplasmic side of this complex (Step 6). Many details in the pathway have yet to be elucidated. Other soluble cellular proteins (e.g. EIF5A) may also play a specific role in the Rev/RRE pathway. It is also not clear what happens once the complex reaches the cytoplasm.
Viral replication inhibited by attacking required steps of Rev function described above has been demonstrated through the use of different classes of Rev mutants (Berger et al. 1991 Virology. 183:630-635). Mutants in each of the Rev functional domains have been described which abolish viral replication. Additionally, a dominant negative form of the Rev protein has been used to inhibit viral replication (Malim et al. 1992. J Exp Med. 176:1197-1201).
The HIV virus particle consists of internal proteins that make up the viral core and two proteins that are part of the lipid envelope that surrounds the core. These proteins are expressed from precursor molecules called Pr55gag and Pr160gagpol for the core proteins and gp160 for the envelope proteins. It is known that expression of these proteins normally requires co-expression of the HIV Rev protein. Without the Rev protein, the mRNAs encoding each of these proteins remains in the nucleus. In order for the Rev protein to work, it is also essential to have an element present in the RNA that binds to Rev. This element is called the RRE.
There is a long felt need in the art for a better method to identify compounds capable of inhibiting HIV replication and Rev function. The present invention satisfies these needs.